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Background: As a FAD (Flavin Adenine Dinucleotide) - dependent histone demethylase discovered in 2004, LSD1 (lysine-specific demethylase 1) was reported to be overexpressed in diverse tumors, regulating target genes transcription associated with cancer development. Hence, LSD1 targeted inhibitors may represent a new insight in anticancer drug discovery. For these reasons, researchers in both the pharmaceutical industry and academia have been actively pursuing LSD1 inhibitors in the quest for new anti-cancer drugs. Objectives: This review summaries patents about LSD1 inhibitors in recent 5 years in the hope of providing a reference for LSD1 researchers to develop new modulators of LSD1 with higher potency and fewer adverse effects. Methods: This review collects LSD1 inhibitors disclosed in patents since 2016. The primary ways of patent searching are Espacenet®, Google Patents, and CNKI. Results: This review covers dozens of patents related to LSD1 inhibitors in recent five years. The compound structures are mainly divided into TCP (Tranylcypromine) derivatives, imidazole derivatives, pyrimidine derivatives, and other natural products and peptides. Meanwhile, the compounds that have entered the clinical phase are also described. Conclusion: Most of the compounds in these patents have been subjected to activity analysis with LSD1 and multi-cell lines, showing good antitumor activity in vitro and in vivo. These patents exhibited the structural diversity of LSD1 inhibitors and the potential of natural products as novel LSD1 inhibitors.

Objectives To explore the long non-coding RNA (lncRNA) expression pattern of congenital lung malformations on a genome-wide scale and investigate their potential biological function in four subtypes of congenital lung malformations. Methods We obtained both lesions and normal lung control tissues from the patients diagnosed with CPAM-I, CPAM-II, ILS, and ILS-CPAM, and underwent lobectomy (i.e., surgical removal of the whole lobe which contains the localized lesion as well as normal lung tissue). Then, we performed lncRNA transcriptome profiling in these tissues by RNA sequencing (RNA-seq). A comprehensive bioinformatics analysis was conducted to characterize the expression profiles and relevant biological functions and for multiple comparisons of lncRNA expression in the different subtypes of congenital lung malformation tissues. Furthermore, the lncRNA-mRNA co-expression network was constructed, and dysregulated mRNAs were functionally analyzed. Finally, gene set enrichment analysis (GSEA) was used to predict the potential molecular mechanism of the identified lncRNAs. Results A total of 5921 lncRNA transcripts were identified between congenital lung malformations tissues and normal lung control tissues. Compared with normal lung control, 481of these expressed lncRNAs were upregulated and 142 were downregulated in CPAM-I, 91 were upregulated and 14 were downregulated in CPAM-II, 39 were upregulated and 38 were downregulated in ILS, and 201 were upregulated and 38 were downregulated in ILS-CPAM. Unsupervised clustering and principal component analysis of the expressed lncRNAs visualized the differences between normal lung control and different subtypes of congenital lung malformations samples. We also confirmed significant differences in the composition of differentially expressed genes (DEGs) and the differentially expressed lncRNAs (DE lncRNAs) between CPAM-I and other subtypes of congenital lung malformations, as well as in normal lung control tissues, and observed enrichment of DEGs in the regulation of the immune system, cell projection organization, and inflammatory pathways. Finally, we identified the lncRNA FLJ26850 might be related to congenital lung malformations via ZNF473. Conclusions Significant differences in lncRNAs expression patterns were observed between different subtypes of congenital lung malformations and normal control. The lncRNA FLJ26850 might be related to congenital lung malformations via ZNF473.

Bitter melon (Momordica charantia) has traditionally been used in the management of diabetes in many countries and territories. However, standardized information on the use of this vegetable as an ...

Zinc finger protein 367 (ZNF367) has been documented as a new cancer-related protein that exerts a pivotal role in the carcinogenesis of multiple cancers. However, whether ZNF367 plays a role in colorectal cancer has not been fully understood. Our data showed that ZNF367 expression was higher in colorectal cancer. Depletion of ZNF367 weakened the proliferative and invasive capabilities of colorectal cancer cells. Up-regulation of ZNF367 enhanced the in vitro malignant features of colorectal cancer cells. Knockdown of ZNF367 impeded the activation of Yes-associated protein (YAP1). Reactivation of YAP1 reversed the ZNF367-knockdown-mediated anticancer effects. Suppression of YAP1 significantly abolished ZNF367-overexpression-induced tumor-promotion effects. Depletion of ZNF367 repressed the tumorigenicity of colorectal cancer cells in vivo. These findings demonstrate that ZNF367 is overexpressed in colorectal cancer and acts as a potential tumor-promoter that contributes to the proliferation and invasion of colorectal cancer by enhancing the activation of YAP1 signaling.

Accurate sweat glucose detection is a promising alternative to invasive finger-prick blood tests, allowing for self-monitoring of blood glucose with good patient compliance. In this study, we have developed a tandem catalytic system, termed as a luminescent "nanochip" (LAON), which was composed of gold nanoparticles (AuNPs) and N-(aminobutyl)-N-(ethylisoluminol) (ABEI)-engineered oxygen-doped carbon nitride (O-g-C3N4), for chemiluminescent detection of sweat glucose. The LAON exhibits dual catalytic activity of glucose oxidase and peroxidase and can not only oxidize glucose to generate H2O2 but catalyze H2O2-mediated luminol chemiluminescence, resulting in sensitive detection of glucose. We identify that the LAON can precisely detect glucose with a detection limit of 0.1 μM, enabling us to measure glucose levels in different biological samples. Particularly, the LAON is capable of sensitively and accurately monitoring dynamic changes in sweat glucose during exercise. Therefore, the LAON provides an alternative approach to supersede invasive blood tests and may improve the management of diabetes mellitus.

The self-assembled phospholipid- or cytosol-associated multienzyme complexes constitute necessary components of the foundation of life. As a proof of concept, metal-coordinated supramolecular nanogels (MCSGs) have been designed, with the self-assembly of di-lysine coordinated iron (Fe(Lys)2)-functionalized peptide gelators on the interface by an in situ amidation-induced protonation process. The monoatomic and highly dispersed active centers of Fe(Lys)2 offered the nanogel mimics with excellent reaction rates due to the high density and nano compartmental structure similar to the natural matrix-associated multienzyme complex. SiO2@MCSGs show both superoxide dismutase (SOD) activity and peroxidase (POD) activity, and the higher activities compared with the activity of free Fe(Lys)2 molecules can be detected. After loading the substrate 2,2′-azinobis-(3-ethylbenzthiazoline-6-sulphonate) (ABTS), SiO2@MCSGsABTS can responsively convert O2− · in the tumor microenvironment into H2O2 intermediates and then tandem catalyzed the oxidization of ABTS for contrast photoacoustic (PA) imaging of tumor by the SOD-POD mimic activity, showing their great potential as the efficient enzymatic agents for pathological theranostics.

Transcription factor-driven cell fate engineering in pluripotency induction, transdifferentiation, and forward reprogramming requires efficiency, speed, and maturity for widespread adoption and clinical translation. Here, we used Oct4, Sox2, Klf4, and c-Myc driven pluripotency reprogramming to evaluate methods for enhancing and tailoring cell fate transitions, through directed evolution with iterative screening of pooled mutant libraries and phenotypic selection. We identified an artificially evolved and enhanced POU factor (ePOU) that substantially outperforms wild-type Oct4 in terms of reprogramming speed and efficiency. In contrast to Oct4, not only can ePOU induce pluripotency with Sox2 alone, but it can also do so in the absence of Sox2 in a three-factor ePOU/Klf4/c-Myc cocktail. Biochemical assays combined with genome-wide analyses showed that ePOU possesses a new preference to dimerize on palindromic DNA elements. Yet, the moderate capacity of Oct4 to function as a pioneer factor, its preference to bind octamer DNA and its capability to dimerize with Sox2 and Sox17 proteins remain unchanged in ePOU. Compared with Oct4, ePOU is thermodynamically stabilized and persists longer in reprogramming cells. In consequence, ePOU: 1) differentially activates several genes hitherto not implicated in reprogramming, 2) reveals an unappreciated role of thyrotropin-releasing hormone signaling, and 3) binds a distinct class of retrotransposons. Collectively, these features enable ePOU to accelerate the establishment of the pluripotency network. This demonstrates that the phenotypic selection of novel factor variants from mammalian cells with desired properties is key to advancing cell fate conversions with artificially evolved biomolecules.

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a potentially curative therapeutic strategy to treat several hematological malignancies and non-hematological malignancies. However, graft-versus-host disease (GVHD) is a frequent and serious transplant-related complication which dramatically restrains the curative effect of allo-HSCT and a significant cause of morbidity and mortality in allogeneic HCT recipients. Effective prevention of GVHD mainly depends on the induction of peripheral immune tolerance. Human leukocyte antigen-G (HLA-G) is a non-classical MHC class I molecule with a strong immunosuppressive function, which plays a prominent role in immune tolerance. HLA-G triggers different reactions depending on the activation state of the immune cells and system. It also exerts a long-term immune tolerance mechanism by inducing regulatory cells. In this present review, we demonstrate the immunomodulatory properties of human leukocyte antigen-G and highlight the role of HLA-G as an immune regulator of GVHD. Furthermore, HLA-G could also serve as a good predictor of GVHD and represent a new therapeutic target for GVHD.

Lipopolysaccharides (LPS) present in the outer leaflet of Gram-negative bacterial outer membranes protect the bacteria from external threats and influence antibiotic permeability as well as immune system recognition. The structure of lipid A, the anchor of an LPS molecule to the outer membrane, can make direct influences on membrane properties. Particularly, in Vibrio cholerae, a Gram-negative bacterium responsible for cholera, a severe diarrheal disease, modifications of lipid A structures grant antibiotic resistance and are a primary factor that led to the current cholera pandemic. However, the difference in structural properties incurred by such modifications has not been fully explored. In this work, five symmetric bilayer systems comprised of distinct lipid A structures of Vibrio cholerae LPS with O1 O-antigen were modeled and simulated to explore influences of different lipid A types on membrane properties. All-atom molecular dynamics simulations reveal that membrane properties such as hydrophobic thickness, acyl chain order parameter, and area per lipid are largely impacted by lipid A modifications due to differences in composition and acyl chain distortions. The modified lipid A is also less negatively charged, which possibly reveals a resistance mechanism to cationic antimicrobial peptide evasion. These findings present a possible explanation for Vibrio cholerae's immune system evasion properties and establish the differences between the lipid A types, which should be of use for any future study of the Gram-negative bacteria.

Objective The aim is to develop a novel noninvasive prenatal testing (NIPT) method that simultaneously performs fetal aneuploidy screening and the detection of de novo and paternally derived mutations. Methods A total of 68 pregnancies, including 26 normal pregnancies, 7 cases with fetal aneuploidies, 7 cases with fetal achondroplasia or thanatophoric dysplasia, 18 cases with fetal skeletal abnormalities, and 10 cases with β‐thalassemia high risk were recruited. Plasma cell‐free DNA was amplified by Targeted And Genome‐wide simultaneous sequencing (TAGs‐seq) to generate around 99% of total reads covering the whole‐genome region and around 1% covering the target genes. The reads on the whole‐genome region were analyzed for fetal aneuploidy using a binary hypothesis T‐score and the reads on target genes were analyzed for point mutations by calculating the minor allelic frequency of loci on FGFR3 and HBB. TAGs‐seq results were compared with conventional NIPT and diagnostic results. Results In each sample, TAGs‐seq generated 44.7–54 million sequencing reads covering the whole‐genome region of 0.1–3× and the target genes of >1000×depth. All cases of fetal aneuploidy and de novo mutations of achondroplasia/thanatophoric dysplasia were identified with high sensitivities and specificities except for one false‐negative paternal mutation of β‐thalassemia. Conclusions TAGs‐seq is a novel NIPT method that combines the fetal aneuploidy screening and the detection of de novo FGFR3 mutations and paternal HBB mutations., Key Points What's already known about this topic? Recently, the improvement of noninvasive prenatal testing (NIPT) allowed the expanded detection of chromosomal copy number variants (CNVs) and monogenic diseases in fetus. However, the expanded detection of CNVs and monogenic disease usually requires separate experiment procedures such as quantitative polymerase chain reaction (qPCR), digital PCR or haplotype‐assisted approaches, which could not be integrated with the screening of chromosome aneuploidy and lack of laboratory efficiency What does this study add? We developed a novel NIPT method that could simultaneously detect fetal aneuploidies, autosomal dominant diseases, and the paternally inherited mutations of autosomal recessive diseases without additional experiments. We validated this method using the plasma samples of pregnant women carrying fetal achondroplasia, thanatophoric dysplasia, and β‐thalassemia, which showed promising results

Broflanilide, a novel insecticide, is classified as a negative allosteric modulator (NAM) of insect γ-aminobutyric acid (GABA) receptors (GABARs) as desmethyl-broflanilide (DMBF) allosterically inhibits the GABA-induced responses. The G277M mutation of the Drosophila melanogaster GABAR subunit has been reported to abolish the inhibitory activity of DMBF. The binding mode of DMBF in insect GABARs needs to be clarified to understand the underlying mechanism of this mutation and to develop novel, efficient NAMs of insect GABARs. Here, we found that a hydrogen bond formed between DMBF and G277 of the D. melanogaster GABAR model might be the key interaction for the antagonism of DMBF by in silico simulations. The volume increase induced by the G277M mutation blocks the entrance of the binding pocket, making it difficult for DMBF to enter the binding pocket and thereby decreasing its activity. The following virtual screening and bioassay results identified a novel NAM candidate of insect GABARs. Overall, we reported a possible binding mode of DMBF in insect GABARs and proposed the insensitivity mechanism of the G277M mutant GABAR to DMBF using molecular simulations. The identified NAM candidates might provide more alternatives or potentials for the design of GABAR-targeting insecticides.

Wound healing is a complex process involved in repairing tissue damage and preventing infection. However, there is a lack of appropriate treatment solutions that can simultaneously promote tissue repair and protect againstbacteria, especially antibiotic-resistant bacteria. In this study, we have developed an injectable hydrogel encapsulating acidic fibroblast growth factor (aFGF) and bacteriophage, termed as ABgel, for combating antibiotic-resistant bacteria and enhancing wound regeneration. ABgel is composed of oxidized sodium alginate (OSA), gelatin and hyaluronic acid (HA), and can rapidly form hydrogel with an elastic modulus of 13 kPa, which mimics the skin tissues. In addition, ABgel can effectively load and stabilize bacteriophage and aFGF, allowing for preventing bacterial infections and improving regeneration of damaged dermal tissues. In vitro studies demonstrate that ABgel exhibits enormous antibacterial activity against antibiotic-resistant E. coli and enhanced angiogenetic activity. Importantly, ABgel can promote skin regeneration and prevent bacterial infections in mice, thereby promoting wound healing process. Therefore, ABgel represents a decent bioactive engineered hydrogel dressing with a broad application potential.

New findings What is the central question of this study? Long non-coding RNAs (lncRNAs) are widely involved in the progression of Hirschsprung's disease (HSCR), but the role of actin filament associated protein 1 antisense RNA1 (AFAP1-AS1), an lncRNA, in HSCR has not been explored before. What is the main finding and its importance? Downregulation of AFAP1-AS1 blocks enteric neural crest stem cell proliferation, differentiation, migration and invasion and promotes the occurrence of HSCR via the miR-195/E2F3 axis, indicating thatAFAP1-AS might be a potential biomarker for HSCR patients. Abstract Long non-coding RNAs (lncRNAs) are involved in several human disorders. Nevertheless, it remains unclear whether they are implicated in the phenotypes of enteric neural crest stem cells (ENCSCs) in Hirschsprung's disease (HSCR). Therefore, we designed this study to explore the pathogenicity of AFAP1-AS1 for HSCR. Microarray analysis and bioinformatic tools were used to screen out the differentially lncRNAs and microRNAs (miRNAs) in patients with HSCR. Small interference RNA transfection was applied to carry out functional experiments in ENCSCs. Cellular activities were detected by cell counting kit-8, 5-ethynyl-2'-deoxyuridine, Transwell assays and flow cytometry. Finally, rescue experiments were performed to examine the cofunction of AFAP1-AS1 and miR-195 and of miR-195 and E2F transcription factor 3 (E2F3). AFAP1-AS1 was reduced in HSCR patients. Meanwhile, knockdown of AFAP1-AS1 reduced the cell migratory and proliferative capacities and facilitated cell apoptosis along with G0/G1 phase arrest. E2F3 was diminished when miR-195 was upregulated, and AFAP1-AS1 inhibition reduced its ability to bind to miR-195. Altogether, AFAP1-AS1 silencing acts as an endogenous RNA by interacting with miR-195 to alter E2F3 expression, thus conferring repressive effects on ENCSC activity and promoting HSCR progression.

Human flap endonuclease 1 (FEN1) is a structure-specific, multifunctional endonuclease essential for DNA replication and repair. Our previous study showed that in response to DNA damage, FEN1 interacts with the PCNA-like Rad9-Rad1-Hus1 complex instead of PCNA to engage in DNA repair activities, such as stalled DNA replication fork repair, and undergoes SUMOylation by SUMO-1. Here, we report that succinylation of FEN1 was stimulated in response to DNA replication fork-stalling agents, such as ultraviolet (UV) irradiation, hydroxyurea, camptothecin, and mitomycin C. K200 is a key succinylation site of FEN1 that is essential for gap endonuclease activity and could be suppressed by methylation and stimulated by phosphorylation to promote SUMO-1 modification. Succinylation at K200 of FEN1 promoted the interaction of FEN1 with the Rad9-Rad1-Hus1 complex to rescue stalled replication forks. Impairment of FEN1 succinylation led to the accumulation of DNA damage and heightened sensitivity to fork-stalling agents. Altogether, our findings suggest an important role of FEN1 succinylation in regulating its roles in DNA replication and repair, thus maintaining genome stability.

In plants, glutathione (GSH) is crucial for the detoxification and tolerance of heavy metals. However, the change characteristics and decisive enzymes involved in GSH metabolism under heavy metal exposure are still unclear. Based on long-term exposure cultivation of spinach and monitoring of the change trends of enzyme activity and GSH contents in response to cadmium (Cd) stress, these issues were clarified. Spinach goes through three statuses in sequence in response to Cd stress, that is, perception status (PS), response status (RS), and new stable status. With the increase in the Cd concentration, the durations of the PS and RS and the time to reach the peaks in the roots were shorter. However, the durations of the PS and the time to reach the peaks in the leaves were longer. The enzyme activities changed significantly in response to diverse Cd stress in RS. γ-glutamyl transpeptidase was vital to the GSH content in roots. Glutathione synthase was important for the GSH content in leaves. The results of this study provide valuable information to find an efficient way to perform GSH adjustments to fulfill the goal of ensuring food safety.

Enterobacterial common antigen (ECA) is a surface glycolipid shared by all members of the Enterobacteriaceae family. In addition to lipopolysaccharides (LPS), ECA is an important component in the outer membrane (OM) of Gram-negative bacteria, making the OM an effective, selective barrier against the permeation of toxic molecules. Previous modeling and simulation studies represented OMs exclusively with LPS in the outer leaflet. In this work, various ECA molecules were first modeled and incorporated into symmetric bilayers with LPS in different ratios, and all-atom molecular dynamics simulations were conducted to investigate the properties of the mixed bilayers mimicking OM outer leaflets. Dynamic and flexible conformational ensembles are sampled for each ECA/LPS system. Incorporation of ECALPS (an LPS core-linked form) and ECAPG (a phosphatidylglycerol-linked form) affects lipid packing and ECA/LPS distributions on the bilayer surface. Hydrophobic thickness and chain order parameter analyses indicate that incorporation of ECAPG makes the acyl chains of LPS more flexible and disordered and thus increases the area per lipid of LPS. The calculated area per lipid of each ECA/LPS provides a good estimate for building more realistic OMs with different ratios of ECA/LPS, which will be useful in order to characterize their interactions with outer membrane proteins in more realistic OMs.

In plants, the spatiotemporal expression of circadian oscillators provides adaptive advantages in diverse species. However, the molecular basis of circadian clock in soybean is not known. In this study, we used soybean hairy roots expression system to monitor endogenous circadian rhythms and the sensitivity of circadian clock to environmental stimuli. We discovered in experiments with constant light and temperature conditions that the promoters of clock genes GmLCLb2 and GmPRR9b1 drive a self-sustained, robust oscillation of about 24-h in soybean hairy roots. Moreover, we demonstrate that circadian clock is entrainable by ambient light/dark or temperature cycles. Specifically, we show that light and cold temperature pulses can induce phase shifts of circadian rhythm, and we found that the magnitude and direction of phase responses depends on the specific time of these two zeitgeber stimuli. We obtained a quadruple mutant lacking the soybean gene GmLCLa1, LCLa2, LCLb1, and LCLb2 using CRISPR, and found that loss-of-function of these four GmLCL orthologs leads to an extreme short-period circadian rhythm and late-flowering phenotype in transgenic soybean. Our study establishes that the morning-phased GmLCLs genes act constitutively to maintain circadian rhythmicity and demonstrates that their absence delays the transition from vegetative growth to reproductive development.

Moraxella catarrhalis (M. catarrhalis) is a pathogenic gram-negative bacterium that causes otitis media and sinusitis in children. Three major serotypes A, B and C are identified to account for approximately 95% of the clinical isolates. Understanding the conformational properties of different serotypes of M. catarrhalis provides insights into antigenic determinants. In this work, all-atom molecular dynamics simulations were conducted for M. catarrhalis lipooligosaccharide (LOS) bilayer systems and oligosaccharides (OS) in water solution to investigate the conformational similarities and differences of three serotypes. For up to 10 neutral monosaccharides in the core part, the conformational ensembles described by the pair-wise root mean square deviation distributions are similar among the three serotypes of either the LOS or OS. At the central β-($1\to4$)-linkage, anti-$\psi$ conformation in conjunction with the gauche-gauche (g−) conformation of the central trisubstituted glucosyl residue is observed as the dominant conformation to sustain the structural characteristics of M. catarrhalis three types, which is further supported by calculated transglycosidic ${}^3{J}_{C,H}\Big({\psi}_H\Big)$ of serotype A in comparison to experimental data. Interestingly, the conformational variability of three serotypes is more restricted for the OS in water solution than that in the LOS bilayer systems. The LOS–LOS interactions in the bilayer systems are responsible for the increased conformational diversity despite of tight packing. Solvent-accessible surface area analysis suggests that a trisaccharide attached to the β-($1\to 6$)-linked sugar in all three serotypes of LOS could be the common epitope and have the possibility to interact with antibodies.

Cotton, the leading natural fiber, is cultivated worldwide, but its production is seriously threatened by pathogens. Accordingly, the selection of resistant cultivars has become a key priority of cotton breeding programs. In this study, a spontaneous mutant with red coloration (S156) and a control cultivar (S78) were used as experimental materials for a comparative analysis. Metabolomic analysis revealed the enrichment of flavonoids in S156 leaves compared with S78 leaves, and transcriptomic analysis revealed the upregulated expression of flavonoid biosynthesis genes in S156 leaves relative to S78 leaves. In addition, the red mutant showed a significantly increase in resistance to Verticillium dahliae, a fungal pathogen that poses a major threat to cotton production. The pathogen invasion process was suppressed in the red cotton cultivar. This study reveals the mechanism underlying the red coloration of S156 cotton and indicates the great potential of red cotton in pathogen- and insect-resistant breeding of cotton.

Apoptotic peptide (kla), which can trigger the mitochondria-mediated apoptotic programmed cell death, has been widely recognized as a potential anticancer agent. However, its therapeutic potential has been significantly impaired by its poor biostability, lack of tumor specificity, and particularly low cellular uptake. Herein, a linear peptide Arg-Trp-d-Arg-Asn-Arg (RWrNR) was identified as an integrin αvβ3 specific ligand with a nanomolar dissociation constant (Kd = 0.95 nM), which can greatly improve kla antitumor activity (IC50 = 8.81 μM) by improving its cellular uptake, compared to the classic integrin-recognition motif c-RGDyK (IC50 = 37.96 μM). Particularly, the RWrNR-kla conjugate can be entrapped in acidic sensitive nanogels (RK/Parg/CMCS-NGs), composed of poly-l-arginine (Parg) and carboxymethyl chitosan (CMCS, pI = 6.8), which can not only carry out controlled release of RWrNR-kla in response to the tumor acidic microenvironment, and consequently enhance its tumor specificity and cell internalization, but also trigger tumor-associated macrophages to generate nitric oxide, leading to enhanced synergistic anticancer efficacy. Importantly, RK/Parg/CMCS-NGs have been proven to effectively activate the apoptosis signaling pathway in vivo and significantly inhibit tumor growth with minimal adverse effects. To summarize, RK/Parg/CMCS-NGs are a promising apoptotic peptide-based therapeutics with enhanced tumor accumulation, cytosolic delivery, and synergistic anticancer effects, thereby holding great potential for the treatment of malignant tumors.

Objective To gain a deeper understanding of the hot topics and future prospects of programmed cell death 1 (PD-1)/programmed cell death ligand 1 (PD-L1) inhibitors treatment of cancer through scientometric analysis of the top-100 most cited clinical trials. Materials and methods We searched the Web of Science Core Collection database from 1980 to June 2019. Two reviewers independently screened the top-100 most cited clinical trials that defined by the National Institutes of Health starting from the most cited article. Title, year of publication, citations, type of cancer, and focused aspects of outcomes were extracted from included clinical trials. VOSviewer software (version 1.6.9) and Excel 2016 were used to do statistical analysis. The evidence mapping was used to present the relationship between cancers, drugs, citations, and outcomes, etc. RESULTS: The top-100 most cited clinical trials published from 2010 to 2018 in nine journals with high impact factor (IF) (IF2018:6.68-70.67), and Lancet Oncology (USA) published the most clinical trials (n = 29, IF2018 = 35.3856). The total number of citations of the top-100 most cited clinical trials was from 59 to 5606. 920 authors from 34 countries and 458 organizations participated in publishing the top-100 most cited clinical trials. The USA (n = 95) and Memorial Sloan-Kettering Cancer Center (n = 31) contributed the most publications. Based on the evidence mapping, there are 25 different types of cancers (e.g. lung cancer, melanoma, and renal cell cancer) and five focused aspects of outcomes (e.g. safety and efficacy). Conclusion The USA was the dominant country. Anti-PD-1/PD-L1 drugs were widely used to treat lung cancer, melanoma, renal cell cancer, and Hodgkin lymphoma. More exploration should be done to explore the use of anti-PD-1/PD-L1 drugs to treat more type of cancers in future research.

Aim: To reconstruct the ancestral sequence of human adenoviral hexon protein by combining sequence variations and structural information. And to provide a candidate hexon protein for developing new adenoviral vector capable of escaping the pre-existing immunity in healthy populations.Methods: The sequences of 74 adenovirus-type strains were used to predict the ancestral sequence of human adenovirus hexon protein using FastML and MEGA software. The three-dimensional structure model was built using homology modeling methods. The immunological features of ancestral loop 1 and loop 2 regions of sequences were tested using protein segments expressed in a prokaryotic expression system and polypeptides synthesized with human serum samples.Results: The tower region of the hexon protein had the highest sequence variability, while the neck and base regions remained constant among different types. The modern strains successfully predicted the common ancestral sequence of the human adenovirus hexon. The positive sera against neutralizing epitopes on the common ancestor of adenoviral hexon were relatively rare among healthy adults.Conclusion: The existing strains inferred the common ancestor of human adenoviruses, with epitopes never observed in the current human strains. The predicted common ancestor hexon is a good prospect in the improvement of adenovirus vectors.

Cell-free fetal DNA fraction (FF) in maternal plasma is a key parameter affecting the performance of noninvasive prenatal testing (NIPT). Accurate quantitation of FF plays a pivotal role in these tests. However, there are few methods that could determine FF with high accuracy using shallow‐depth whole‐genome sequencing data. In this study, we hypothesized that the actual FF in maternal plasma should be proportional to the discrepancy rate between the observed genotypes and inferred genotypes based on the linkage disequilibrium rule in certain polymorphism sites. Based on this hypothesis, we developed a method named Linkage Disequilibrium information-based cell-free Fetal DNA Fraction (LDFF) to accurately quantify FF in maternal plasma. This method achieves a high performance and outperforms existing methods in the fetal DNA fraction estimation. As LDFF is a gender-independent method and developed on shallow-depth samples, it can be easily incorporated into routine NIPT test and may enhance the current NIPT performance.

Ya Gao,1,* Weiru Li,1,* Xiaoyin Bu,1,* Ying Xu,1 Shengchun Cai,1 Jinman Zhong,1 Meixue Du,1 Haitao Sun,2 Liping Huang,3 Yongjian He,4 Xiumei Hu,4 Qifa Liu,1 Hua Jin,1 Qian Wang,2,4 Baohong Ping5 1Department of Hematology, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, People’s Republic of China; 2Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, 510280, People’s Republic of China; 3Department of Obstetrics and Gynecology, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, People’s Republic of China; 4Department of Laboratory Medicine, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, People’s Republic of China; 5Department of Hematology, Huiqiao Medical Center, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, People’s Republic of China*These authors contributed equally to this workCorrespondence: Baohong PingDepartment of Hematology, Huiqiao Medical Center, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, People’s Republic of ChinaTel +8615625042109Fax +862061642231Email nfyypingbaohong66@163.comQian WangDepartment of Laboratory Medicine, Nanfang Hospital, Southern Medical University, Guangzhou, 510515, People’s Republic of ChinaTel +8613822251298Fax +862061643018Email wangqian@smu.edu.cnBackground: Acute graft versus host disease (aGVHD) remains a leading cause of transplant-related mortality following allogeneic haematopoietic cell transplantation (allo-HCT). Human amniotic mesenchymal stem cells (hAMSCs) are a novel mesenchymal stem cells (MSCs), which have stronger proliferation and immunomodulatory ability compared with bone marrow mesenchymal stem cells (BM-MSCs). Besides, as the amniotic membrane is often treated as medical waste after delivery, hAMSCs can be obtained conveniently and noninvasively. The aim of this study was to explore the therapeutic efficacy and underlying mechanisms of hAMSCs transplantation for the humanized aGVHD mouse model.Methods: We established a humanized aGVHD mouse model by transplanting human peripheral blood mononuclear cells (PBMCs) into NOD-PrkdcscidIL2rγnull (NPG) mice, human amniotic membrane collected from discarded placenta of healthy pregnant women after delivery and hAMSCs were extracted from amniotic membrane and expanded in vitro. Mice were divided into untreated group (Control), aGVHD group (aGVHD), and hAMSCs treatment group (aGVHD+hAMSCs), the hAMSCs labeled with GFP were administered to aGVHD mice to explore the homing ability of hAMSCs. T effector and regulatory T cells (Tregs) levels and cytokines of each group in target organs were detected by flow cytometry and cytometric bead array (CBA), respectively.Results: We successfully established a humanized aGVHD mouse model using NPG mice. The hAMSCs have the ability to inhibit aGVHD in this mouse model through reduced villous blunting and lymphocyte infiltration of the gut while reducing inflammatory edema, tissue destruction and lymphocyte infiltration into the parenchyma of the liver and lung. hAMSCs suppressed CD3+CD4+ T and CD3+CD8+ T cell expression and increased the proportion of Tregs, and besides, hAMSCs can reduce the levels of IL-17A, INF-γ, and TNF in aGVHD target organs.Conclusion: The NPG murine environment was capable of activating human T cells to produce aGVHD pathology to mimic aGVHD as in humans. The hAMSCs controlled aGVHD by decreasing inflammatory cytokine secretion within target organs by modulating the balance of Tregs and T effector cells in humanized mice.Keywords: acute graft versus host disease, humanized mouse model, NPG mice, amniotic mesenchymal stem cells, immunomodulatory

Long non-coding RNAs (lncRNAs) and microRNAs (miRNAs) are essential for the progression of tumors, including cutaneous squamous cell carcinoma (CSCC). The present study aimed to examine the competing endogenous RNA (ceRNA) network in CSCC. Differentially expressed genes in CSCC were analyzed using the GSE66359 microarray data set, and the upstream miRNAs and lncRNAs were predicted using online database analysis (TargetScan 7.1, mirDIP 4.1, miRSearch V3.0, miRDB and RNA22 2.0) and were verified in clinical tissues. RNA pull-down and dual luciferase reporter gene assays were used to verify the targeting relationships among lncRNA human histocompatibility leukocyte antigen complex P5 (HCP5), miR-138-5p and enhancer of zeste homolog 2 (EZH2). Cell lines with a high and low HCP5 expression were screened, and a pcDNA-3.1-HCP5 overexpression vector, small interfering RNA against HCP5, miR-138-5p mimics and miR-138-5p inhibitors were transfected into the CSCC cells. Cell viability, invasion, migration, apoptotic rate and autophagy were evaluated. The effects of HCP5 on autophagy and apoptosis of CSCC cells were verified in vivo using Ki67 and TUNEL staining. EZH2 was demonstrated to be upregulated in CSCC cells. miR-138-5p target sequences were identified in HCP5 and EZH2. HCP5 was revealed to function as a putative ceRNA of miR-138-5p to positively regulate EZH2, and EZH2 was shown to regulate autophagy and apoptosis of CSCC cells through the STAT3/VEGFR2 pathway. HCP5 overexpression decreased miR-138-5p levels, increased EZH2 levels and promoted cell malignant behaviors and autophagy but decreased the apoptosis rate. These trends were opposite when HCP5 was silenced. In conclusion, HCP5 may competitively bind to miR-138-5p to regulate EZH2 in CSCC cells, promoting autophagy and reducing apoptosis through the STAT3/VEGFR2 pathway. This study may provide a new perspective for understanding the molecular mechanism and treatment of CSCC.

Background Subtilisin QK is a serine protease in the subtilisin family, and is fermented by Bacillus subtilis QK02. The fibrinolytic activity of subtilisin QK was measured by detecting low molecular weight degradation products using a spectrophotometric method developed by Japan Bio Science Laboratory Co., Ltd. Subtilisin QK powder can maintain its fibrinolytic activity for more than 24 months when it is stored at room temperature and protected from light. Our previous results showed that subtlisin QK directly degraded cross-linked fibrins in the fibrin plate assay and effectively inhibited thrombosis in the mouse thrombus model. The aim of this study was to determine the acute toxicity, potential subchronic toxicity, and safety pharmacology of subtilisin QK in Sprague–Dawley (SD) rats. Methods In the acute toxicity study, a single oral dose of 100,000 FU/kg was administered to 10 female and 10 male SD rats. In the 28-day subchronic toxicity, 60 female and 60 male SD rats were randomly assigned to four experimental groups (daily oral dose of 0, 2500, 7500 and 25,000 FU/kg). In the safety pharmacology study, 20 female and 20 male SD rats were randomly assigned to four experimental groups (single oral dose of 0, 500, 1500 and 5000 FU/kg). Results No death occurred and no adverse effects were observed in the acute toxicity study at a dose of 100,000 FU/kg. In the 28-day subchronic toxicity study, several hematological and blood biochemical parameters showed increases or decreases; however, due to the lack of a dose–response relationship, these differences were considered unrelated to treatment. In the safety pharmacology study, no adverse effects were observed on the central nervous of SD rats post-administration up to a dose of 5000 FU/kg subtilisin QK. Conclusion The results showed that oral consumption of subtilisin QK is of low toxicological concern. No adverse effects were observed at doses of 2500, 7500, and 25,000 FU/kg in the 28-day subchronic toxicity, and the no-observed-adverse-effect level (NOAEL) of subtilisin QK was 25,000 FU/kg.

Mastitis caused by Escherichia coli (E. coli) remains a threat to dairy animals and impacts animal welfare and causes great economic loss. Furthermore, antibiotic resistance and the lagged development of novel antibacterial drugs greatly challenge the livestock industry. Phage therapy has regained attention. In this study, three lytic phages, termed vB_EcoM_SYGD1 (SYGD1), vB_EcoP_SYGE1 (SYGE1), and vB_EcoM_SYGMH1 (SYGMH1), were isolated from sewage of dairy farm. The three phages showed a broad host range and high bacteriolytic efficiency against E. coli from different sources. Genome sequence and transmission electron microscope analysis revealed that SYGD1 and SYGMH1 belong to the Myoviridae, and SYGE1 belong to the Autographiviridae of the order Caudovirales. All three phages remained stable under a wide range of temperatures or pH and were almost unaffected in chloroform. Specially, a mastitis infected cow model, which challenged by a drug resistant E. coli, was used to evaluate the efficacy of phages. The results showed that the cocktails consists of three phages significantly reduced the number of bacteria, somatic cells, and inflammatory factors, alleviated the symptoms of mastitis in cattle, and achieved the same effect as antibiotic treatment. Overall, our study demonstrated that phage cocktail may be a promising alternative therapy against mastitis caused by drug resistant E. coli.

Pseudomonas aeruginosa is a Gram-negative pathogen that causes a variety of infections in humans and animals. Due to the inappropriate use of antibiotics, multi-drug resistant (MDR) P. aeruginosa strains have emerged and are prevailing. In recent years, cow mastitis caused by MDR P. aeruginosa has attracted attention. In this study, a microbial community analysis revealed that P. aeruginosa could be a cause of pathogen-induced cow mastitis. Five MDR P. aeruginosa strains were isolated from milk diagnosed as mastitis positive. To seek an alternative antibacterial agent against MDR, P. aeruginosa, a lytic phage, designated vB_PaeS_PAJD-1 (PAJD-1), was isolated from dairy farm sewage. PAJD-1 was morphologically classified as Siphoviridae and was estimated to be about 57.9 kb. Phage PAJD-1 showed broad host ranges and a strong lytic ability. A one-step growth curve analysis showed a relatively short latency period (20 min) and a relatively high burst size (223 PFU per infected cell). Phage PAJD-1 remained stable over wide temperature and pH ranges. Intramammary-administered PAJD-1 reduced bacterial concentrations and repaired mammary glands in mice with mastitis induced by MDR P. aeruginosa. Furthermore, the cell wall hydrolase (termed endolysin) from phage PAJD-1 exhibited a strong bacteriolytic and a wide antibacterial spectrum against MDR P. aeruginosa. These findings present phage PAJD-1 as a candidate for phagotherapy against MDR P. aeruginosa infection.

Background: Acute graft versus host disease (aGVHD) remains a leading cause of transplant-related mortality following allogeneic haematopoietic cell transplantation(allo-HCT). Although previous studies indicated that mesenchymal stem cells (MSCs) may be a salvage therapeutic agent for aGVHD, the mechanism is not yet fully clear. Human amniotic mesenchymal stem cells (hAMSCs) is a novel MSCs, compared with bone marrow mesenchymal stem cells, it has the advantage of being non-invasive, and also has stronger proliferation ability than that of BM-MSCs and equivalent immune regulation ability as BM-MSCs. The aim of this study was to explore the therapeutic efficacy and underlying mechanisms of human amniotic mesenchymal stem cells transplantation for the humanized aGVHD mouse model.Methods: We established a humanized aGVHD mouse model by transplanting human peripheral blood mononuclear cells (PBMCs) into NOD-PrkdcscidIL2rγnull (NPG) mice, hAMSCs collected from discarded placenta of healthy pregnant women after delivery. Mice were divided into control group (untreated), aGVHD group, and hAMSCs treatment group, the hAMSCs labeled with GFP were administered to aGVHD mice to explore the homing ability of hAMSCs. T effector and Treg cell levels and cytokines of each group in target organs were detected by flow cytometry and cytometric bead array (CBA) respectively.Results: We successfully established a humanized aGVHD mouse model using NPG mice. The hAMSCs have the ability to inhibit aGVHD in this mouse model through reduced villous blunting and lymphocyte infiltration into the lamina propria of the gut while reducing vascular endothelialitis and lymphocyte infiltration into the parenchyma of the liver and lung. hAMSCs suppressed xenogenesis CD3+CD4+ T and CD3+CD8+ T cell expression and increased the proportion of Treg cells, and besides, hAMSCs can reduce the levels of IL-17A, INF-γ, TNF, and IL-2 in aGVHD target organs.Conclusions: The NPG murine environment was capable of activating human T cells to produce aGVHD pathology to mimic aGVHD as in humans. The hAMSCs controlled aGVHD by decreasing inflammatory cytokine secretion within target organs by modulating the balance of Treg and T effector cells in humanized mice.

Inflammatory bowel disease (IBD) is a chronic intestinal disorder threatening human health. Di-peptide alanyl-glutamine (Ala-Gln) has various beneficial effects on gut health. However, its role and functional mechanism in treating IBD are still not clear. Therefore, the protective effects of Ala-Gln and glutamine (Gln) on dextran sulfate sodium- (DSS-) induced colitic mice were investigated in this study. The results showed that oral supplementation of Ala-Gln or Gln significantly attenuated the colitis symptoms in mice, including body weight loss, colon length, disease activity index, histological scores, and tissue apoptosis. The concentrations of interleukin- (IL-) 1β, IL-6, tumor necrosis factor-α, and myeloperoxidase were significantly decreased, while the concentrations of immunoglobulins (IgA, IgG, and IgM) and superoxide dismutase were significantly increased by Ala-Gln or Gln supplementation. The expression of occludin and peptide transporter 1 (PepT1) was significantly increased by Ala-Gln or Gln. Interestingly, Ala-Gln had better beneficial effects than Gln in alleviating colitis. In addition, 16S rDNA sequencing showed that the DSS-induced shifts of the microbiome (community diversity, evenness, richness, and composition) in the mouse colon were restored by Gln and Ala-Gln, including Lactobacillus, Bacteroides_acidifaciens, Bacteroidales, Firmicutes, Clostridia, Helicobacter, and Bacteroides. Correspondingly, the functions of the microflora metabolism pathways were also rescued by Ala-Gln, including fatty acid metabolism, membrane transporters, infectious diseases, and immune system. In conclusion, the results revealed that Ala-Gln can prevent colitis through PepT1, enhancing the intestinal barrier and modulating gut microbiota and microflora metabolites.

This is a protocol for a Cochrane Review (intervention). The objectives are as follows: To evaluate the effectiveness and safety of tamoxifen and aromatase inhibitors for women with recurrent high‐grade serous tubo‐ovarian cancer where further conventional chemotherapy or molecular‐targeted cytotoxic drugs are not available or would not be tolerated.

Ruminants are mostly herbivorous animals that employ rumen fermentation for the digestion of feed materials, including dairy cows. Ruminants consume plant fibre as their regular diet, but lack the machinery for their digestion. For this reason, ruminants maintain a symbiotic relation with microorganisms that are capable of producing enzymes to degrade plant polymers. Various species of microflora including bacteria, protozoa, fungi, archaea, and bacteriophages are hosted at distinct concentrations for accomplishing complete digestion. The ingested feed is digested at a defined stratum. The polysaccharic plant fibrils are degraded by cellulolytic bacteria, and the substrate formed is acted upon by other bacteria. This sequential degradative mechanism forms the base of complete digestion as well as harvesting energy from the ingested feed. The composition of microbiota readily gets tuned to the changes in the feed habits of the dairy cow. The overall energy production as well as digestion is decided by the intactness of the resident communal flora. Disturbances in the homogeneity gastrointestinal microflora has severe effects on the digestive system and various other organs. This disharmony in communal relationship also causes various metabolic disorders. The dominance of methanogens sometimes lead to bloating, and high sugar feed culminates in ruminal acidosis. Likewise, disruptive microfloral constitution also ignites reticuloperitonitis, ulcers, diarrhoea, etc. The role of symbiotic microflora in the occurrence and progress of a few important metabolic diseases are discussed in this review. Future studies in multiomics provides platform to determine the physiological and phenotypical upgradation of dairy cow for milk production.

This study elucidated copper (Cu) transport and trafficking mechanisms at the cellular interface using the Cu-resistant strain Klebsiella oxytoca CAV 1374. The optimum conditions for biosorption were determined by investigating uptake rates due to initial pH, initial Cu concentrations, and maximum tolerated concentrations (MTC) of Cu. Cellular fraction analysis and depth-profiling XPS were used to comprehensively evaluate the spatial-temporal distribution of Cu on cellular interfaces during biosorption. Potential uptake mechanisms were then further examined by biosorption kinetics analyses, ion exchange experiments, FTIR analysis, and K+ channel blocking experiments. The results indicated that Cu was primarily absorbed by extracellular polymeric substances through chemical interactions and little Cu penetrated inside cells under low Cu stress conditions (≤20 mg/L). In contrast, an intracellular rate-controlling physical interaction was predominant under high Cu stress conditions (≥30 mg/L). Further, Cu2+ could be bound by functional groups, followed first by replacement of Ca2+ at the cell surface. Subsequently, some of the Cu2+ in cell walls was reduced to Cu+, and only Cu+ could then penetrate into cell membranes. These results indicate that strain Klebsiella oxytoca CAV 1374 is a suitable biosorbent agent for Cu removal and can provide critical insights into Cu-uptake mechanisms of microorganisms.

Ubiquitin-conjugating enzyme E2 M (UBE2M) and ubiquitin-conjugating enzyme E2 F (UBE2F) are the two NEDD8-conjugating enzymes of the neddylation pathway that take part in posttranslational modification and change the activity of target proteins. The activity of E2 enzymes requires both a 26-residue N-terminal docking peptide and a conserved E2 catalytic core domain, which is the basis for the transfer of neural precursor cell-expressed developmentally downregulated 8 (NEDD8). By recruiting E3 ligases and targeting cullin and non-cullin substrates, UBE2M and UBE2F play diverse biological roles. Currently, there are several inhibitors that target the UBE2M-defective in cullin neddylation protein 1 (DCN1) interaction to treat cancer. As described above, this review provides insights into the mechanism of UBE2M and UBE2F and emphasizes these two E2 enzymes as appealing therapeutic targets for the treatment of cancers.

BRCA1 is a tumor suppressor that has been found to be involved DNA synthesis during cell replication. In a recent study, the single nucleotide polymorphism (SNP), rs799917, in BRCA1 was found to be associated with the development and progression of various types of tumor. In the present study, the association between rs799917 and susceptibility to lung cancer was evaluated in a Han Chinese population in the Liaoning Province of China. The BRCA1 rs799917 genotypes (C/C, C/T and T/T) were analyzed using TaqMan quantitative PCR in 682 patients with lung cancer and 694 healthy controls, and the results were analyzed using a Student's t-test, a χ(2) test and logistic regression analysis. Individuals carrying the C/T or T/T genotype had a lower risk of lung cancer compared with those carrying the C/C genotype [odds ratio (OR), 0.741; P=0.021; and OR, 0.610; P=0.011, respectively). The C/T + T/T genotype group had an even lower risk (OR, 0.709; P=0.005) compared with that in the C/C genotype group. In the stratified analyses of non-smokers, individuals with the C/T or T/T genotype had a lower risk of developing lung cancer compared with that in those carrying the C/C genotype (OR, 0.681; P=0.013; and OR, 0.569; P=0.021, respectively). The stratified analyses of the BRCA1 rs799917 polymorphism based on pathological type, chemotherapy and radiotherapy, showed that in the squamous cell carcinoma, non-chemotherapy and non-radiotherapy subgroups, individuals with the T/T genotype had a lower risk of lung cancer compared with that in those carrying the C/C genotype (OR, 0.454; P=0.007; OR, 0.485; P=0.002; and OR, 0.599; P=0.020, respectively). In conclusion, the T allele of the rs799917 SNP in BRCA1 was associated with a lower risk of lung cancer in the ethnic Han Chinese population in Liaoning Province and may represent a protective factor against lung cancer.

An accurate measurement of sweat glucose is a promising alternative to invasive finger prick blood test, and may provide effective self-monitoring of blood glucose with good patient compliance. Herein, an interrelated catalytic enzyme system has been developed, termed as CataFlower, which is composed of nanoflower MoS2 (peroxidase) decorated with GOx (glucose oxidase) and MnO2 (oxygen generator), and exhibits synergistic oxidative capability for sensitively monitoring sweat glucose. CataFlower can not only generate oxygen in situ to maximize GOx activity, but promote peroxidase-triggered H2O2 oxidation of methylene blue, resulting in sensitive colorimetric detection of glucose. We identify that CataFlower can precisely detect glucose with a detection limit of 10 μM, allowing for measuring glucose levels in different biological samples, such as blood and urine. Particularly, CataFlower is capable of monitoring dynamic changes in sweat glucose with high sensitivity and accuracy during exercise. Therefore, CataFlower provides a stepping stone to eliminate invasive blood tests, significantly improving the diagnosis and management of diabetes mellitus.

Primary cilia are compartmentalized sensory organelles present on the majority of neurons in the mammalian brain throughout adulthood. Recent evidence suggests that cilia regulate multiple aspects of neuronal development, including the maintenance of neuronal connectivity. However, whether ciliary signals can dynamically modulate postnatal circuit excitability is unknown. Here we show that acute cell-autonomous knockdown of ciliary signaling rapidly strengthens glutamatergic inputs onto cultured rat neocortical pyramidal neurons and increases spontaneous firing. This increased excitability occurs without changes to passive neuronal properties or intrinsic excitability. Further, the neuropeptide receptor somatostatin receptor 3 (SSTR3) is localized nearly exclusively to excitatory neuron cilia both in vivo and in culture, and pharmacological manipulation of SSTR3 signaling bidirectionally modulates excitatory synaptic inputs onto these neurons. Our results indicate that ciliary neuropeptidergic signaling dynamically modulates excitatory synapses and suggest that defects in this regulation may underlie a subset of behavioral and cognitive disorders associated with ciliopathies.

Centriole duplication is tightly controlled to occur once per cell cycle, and disruption of this synchrony causes centriole amplification, which is frequently observed in many cancers. Our previous work showed that nuclear distribution gene C (NudC)-like protein 2 (NudCL2) localizes to centrosomes; however, little is known about the role of NudCL2 in the regulation of centrosome function. Here, we find that NudCL2 is required for accurate centriole duplication by stabilizing the E3 ligase HECT domain and RCC1-like domain-containing protein 2 (HERC2). Knockout (KO) of NudCL2 using CRISPR/Cas9-based genome editing or depletion of NudCL2 using small interfering RNA causes significant centriole amplification. Overexpression of NudCL2 significantly suppresses hydroxyurea-induced centriole overduplication. Quantitative proteomic analysis reveals that HERC2 is downregulated in NudCL2 KO cells. NudCL2 is shown to interact with and stabilize HERC2. Depletion of HERC2 leads to the similar defects to that in NudCL2-downregulated cells, and ectopic expression of HERC2 effectively rescues the centriole amplification caused by the loss of NudCL2, whereas the defects induced by HERC2 depletion cannot be reversed by exogenous expression of NudCL2. Either loss of NudCL2 or depletion of HERC2 leads to the accumulation of ubiquitin-specific peptidase 33 (USP33), a centrosomal protein that positively regulates centriole duplication. Moreover, knockdown of USP33 reverses centriole amplification in both NudCL2 KO and HERC2-depleted cells. Taken together, our data suggest that NudCL2 plays an important role in maintaining the fidelity of centriole duplication by stabilizing HERC2 to control USP33 protein levels, providing a previously undescribed mechanism restraining centriole amplification.

The cutaneous squamous cell carcinoma (cSCC) originates from epithelial stem cells through the dysregulation of self-renewal and differentiation. Recent studies have identified methyltransferase-like 3 (METTL3)-mediated N6-methyladenosine (m6A) modification as key regulator of fate of stem cells. However, little is known about the functional importance of METTL3 in cSCC. Here, Western blot and immunohistochemistry were used to investigate the METTL3 levels in cSCC tissues. Functional experiments including surface marker detection, Brdu incorporation assay, colony forming assay, m6A dot blot and tumor xenograft assay were performed to investigate the properties in cSCC cell lines after METTL3 knock down. The expression of METTL3 was up-regulated in cSCC samples. METTL3 knock down impaired cSCC cell stem-like properties, including colony forming ability in vitro and tumorigenicity in vivo. Furthermore, METTL3 knock down and methylation inhibitor cycloleucine could decrease the m6A levels and the expression of ΔNp63 in cSCC. Exogenous expression of ΔNp63 partially restored the cell proliferation of METTL3-knockdown cSCC cells. Therefore, our data indicated the m6A methyltransferases METTL3 as a critical gene in regulating tumorigeneis of cSCC.

Objectives To identify the cooperation of authors, countries, and institutions and explore the hot topics and future prospects regarding programmed cell death protein 1 (PD-1) and programmed cell death 1 ligand 1 (PD-L1) research. Materials and methods Publications on PD-1 and PD-L1 research were retrieved from the Web of Science Core Collection. Bibliometric analyses were performed using VOSviewer 1.6.9, HistCite 2.1, and CiteSpace V software. Network maps were generated to evaluate the collaborations between different authors, countries, and institutions. Results A total of 7359 articles related to PD-1 and PD-L1 research in the field of cancer were identified. We observed rapid growth in the number of publications since 2014. Oncoimmunology (326, 4.43%) had the highest number of publications, while N Engl J Med (18,828 co-citations, 6.30%) was the most co-cited journal. Hodi FS and Wolchok JD were key researchers. There were active collaborations among the top authors. The USA was the leading contributor in this field with 3496 publications. Active cooperations between countries and between institutions were observed. The main hot topics included matters related to the efficacy and prognosis of various tumors after treatment with PD-1 and/or PD-L1 inhibitors, as well as the activation or expression of important immunological factors targeted at tumors. Conclusion PD-1 and PD-L1 studies have significantly increased after 2014. The USA contributed the most publications. There were active cooperations between authors, countries, and institutions. Further research should expand and develop new topics such as those likely to boost therapeutic strategies for cancer.

A continuous fibroblast-like cell line, TMF (turbot muscle fibroblasts), was established from juvenile turbot Scophthalmus maximus muscle with the method of trypsin digestion. It has been subcultured more than 60 passages for over 150 days. The TMF cells were cultured in L-15 medium supplemented with HEPES, fetal bovine serum (FBS), GlutaMAX, and basic fibroblast growth factor (bFGF). The optimal temperature for TMF culture was 24 °C. TMF cells were predominantly composed of fibroblastic-like cells, and the transcription factor 4 (TCF-4) was highly expressed in TMF cells. Chromosome analysis revealed that it had a diploid chromosome number of 2n = 44. The transfection efficiency achieved 54.95 ± 6.59%, and the cell mortality rate was about 8.70% when transfected with the nucleofection method. Meanwhile, the TMF cells showed a sensitive response to amino acid levels and activation target of rapamycin (TOR) signaling pathway. These results indicate that TMF was a potential tool to explore the signal transduction of teleost in vitro.

Microneedles are primarily designed for enhancing transdermal drug delivery in a minimally invasive manner. In particular, coated microneedles have attracted increasing attention due to the resulting mode of rapid and highly efficient drug administration. In this study, we developed a novel gelatin/sucrose film-coated poly(ethylene glycol)diacrylate (PEGDA) microneedle patch, which can effectively improve the skin permeability of therapeutic drugs and realize convenient and efficient drug administration. A simple and reliable technique was proposed to produce uniform film coatings on microneedles at room temperature. After introducing a prepolymer solution into the mold cavity, a film-coated microneedle patch was fabricated via a photo-induced polymerization process. Four categories of molecular models, including rhodamine B (RhB), bovine serum albumin (BSA), doxorubicin (DOX) and indocyanine green (ICG), were encapsulated into the gelatin/sucrose film to evaluate their transdermal delivery and therapeutic effects both in vitro and in vivo. The presented methodology represents a facile technique for rapid formation of film-coated microneedles for efficient transdermal delivery of small molecular drugs and large proteins.

HIV reverse transcriptase (RT)-associated ribonuclease H (RNase H) plays an important role in HIV multiplication and represents a novel anti-HIV target. Recently, a novel series of 3-hydroxypyrimidine-2,4-dione (HPD) derivatives have been reported as potent inhibitors of HIV RT-associated RNase H; they also exhibit antiviral activities, with EC50 values in the low micromolar range. To better understand their structure–activity relationships and mechanisms of action, an integrated computational study, including molecular docking, molecular dynamics (MD), three-dimensional quantitative structure–activity relationship (3D-QSAR), and pharmacophore modeling, was performed on these HPDs. Ninety-three HPDs were firstly docked into the RNase H active site; then, MD simulations were performed to validate the accuracy of the docking results. The comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) methods were used to generate 3D-QSAR models. Structure-based CoMFA (q2 = 0.908, R2 = 0.978, rpred2 = 0.949) and CoMSIA (q2 = 0.908, R2 = 0.960, rpred2 = 0.919) models and docking-based CoMFA (q2 = 0.832, R2 = 0.985, rpred2 = 0.967) and CoMSIA (q2 = 0.927, R2 = 0.990, rpred2 = 0.977) models were constructed and exhibited excellent predictive ability. The generated pharmacophore model provided deep insight into the pharmacological structural characteristics of the HPDs. Nine virtually screened compounds and six newly designed compounds based on the pharmacophore and 3D-QSAR models are potential leads of HIV RNase H inhibitors. These results may provide important information for the design and development of potent and novel HIV RNase H inhibitors.

The lactic acid bacterial diversity of wheat sourdoughs collected in Ya'an city, Sichuan province was investigated by culture-dependent methods combined with 16S rRNA gene sequencing, and the exopolysaccharide (EPS) production characteristics of ropy lactic acid bacteria (LAB) strains from these sourdoughs were evaluated. Two hundred nineteen LAB strains were isolated, and genotypic characterization indicated that the isolated LAB strains included Lactobacillus plantarum (L. plantarum), L. pantheris, Leuconostoc citreum (Leu. citreum), Weissella viridescens, Leu. pseudomesenteroides, Lactococcus lactis (L. lactis), L. raffinolactis, and Leu. mesenteroides. All the isolates were found to harbor genes coding for EPS, and the yield of EPS produced by “ropy” LAB strains ranged between 2.3 and 229.5 mg/g wet cell weight. The present study enhances our knowledge about LAB diversity in Ya'an sourdoughs and provides information about the EPS-producing LAB strains in sourdough samples, which might be favorable for the application of specific LAB strains in sourdough technology.

Classical swine fever (CSF), caused by classical swine fever virus (CSFV), is a highly contagious disease of swine with high morbidity and mortality that negatively affects the pig industry worldwide, in particular in China. Soon after the endocytosis of CSFV, the virus makes full use of the components of host cells to complete its life cycle. The endocytosis sorting complex required for transport (ESCRT) system is a central molecular machine for membrane protein sorting and scission in eukaryotic cells that plays an essential role in many physiological metabolic processes, including invasion and egress of envelope viruses. However, the molecular mechanism that ESCRT uses to regulate the replication of CSFV is unknown. In this study, we demonstrated that the ESCRT-I complex Tsg101 protein participates in clathrin-mediated endocytosis of CSFV and is also involved in CSFV trafficking. Tsg101 assists the virus in entering the host cell through the late endosome (Rab7 and Rab9) and finally reaching the lysosome (Lamp-1). Interestingly, Tsg101 is also involved in the viral replication process by interacting with nonstructural proteins 4B and 5B of CSFV. Finally, confocal microscopy showed that the replication complex of Tsg101 and double-stranded RNA (dsRNA) or NS4B and NS5B protein was close to the endoplasmic reticulum (ER), not the Golgi, in the cytoplasm. Collectively, our finding highlights that Tsg101 regulates the process of CSFV entry and replication, indicating that the ESCRT plays an important role in the life cycle of CSFV. Thus, ESCRT molecules could serve as therapeutic targets to combat CSFV infection. IMPORTANCE CSF, caused by CSFV, is a World Organization for Animal Health (OIE) notifiable disease and causes significant financial losses to the pig industry globally. The ESCRT machinery plays an important regulatory role in several members of the genera Flavivirus and Hepacivirus within the family Flaviviridae, such as hepatitis C virus, Japanese encephalitis virus, and dengue virus. Previous reports have shown that assembling and budding of these viruses require ESCRT. However, the role of ESCRT in Pestivirus infection remains to be elucidated. We determined the molecular mechanisms of the regulation of CSFV infection by the major subunit Tsg101 of ESCRT-I. Interestingly, Tsg101 plays an essential regulatory role in both clathrin-mediated endocytosis and genome replication of CSFV. Overall, the results of this study provide further insights into the molecular function of ESCRT-I complex protein Tsg101 during CSFV infection, which may serve as a molecular target for pestivirus inhibitors.

Although previous reports have shown that Curcumin inhibits many viruses, including some important members of different genera of Flaviviridae family (Japanese encephalitis virus, dengue virus and hepatitis C virus), the antiviral activity of curcumin against Classical swine fever virus (CSFV), which belongs to Pestivirus genus, is still unclear. In this study, we found that curcumin inhibited CSFV replication in a dose-dependent manner, but had no effect on virus adsorption and entry. Furthermore, the results showed that curcumin inhibited the expression of FASN, one of the key enzymes of fatty acid synthesis pathway, thereby, causing the reduction of the production of LDs upon infection. To this end, we detected transcription factor 6 (ATF6), the key factor of regulating lipid metabolism along with other related molecules (CHOP and GPR78) and found that curcumin significantly impaired the gene synthesis of ATF6, while CSFV infection promoted ATF6 expression. Therefore, it is confirmed that curcumin inhibited CSFV replication by interfere lipid metabolism. In addition, our subsequent studies found that curcumin played an antiviral role by promoting the innate immune independent of NF-κB signaling pathway. Taken together, our finding highlights that curcumin is a potential candidate drug against CSFV for controlling CSF.

Streptococcus suis (S. suis) serotype 2 (SS2) is the causative agent of swine streptococcosis and can cause severe diseases in both pigs and humans. Although the traditional inactive vaccine can protect pigs from SS2 infection, novel vaccine candidates are needed to overcome its shortcomings. Three infection-associated proteins in S. suis—muramidase-released protein (MRP), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and DLD, a novel putative dihydrolipoamide dehydrogenase—have been previously identified by immunoproteomic assays. In this study, the effective immune protection of the recombinant trivalent protein GAPDH-MRP-DLD (JointS) against SS2, SS7, and SS9 was determined in zebrafish. To improve the immune efficacy of JointS, monophosphoryl lipid A (MPLA) as a TLR4 agonist adjuvant, which induces a strong innate immune response in the immune cells of mice and pigs, was combined with JointS to immunize the mice. The results showed that immunized mice could induce the production of a high titer of anti-S. suis antibodies, as a result, 100% of mice survived after SS2 infection. Furthermore, JointS provides good protection against virulent SS2 strain infections in piglets. Given the above, there is potential to develop JointS as a novel subunit vaccine for piglets to prevent infection by SS2 and other S. suis serotypes.

SummaryThe cavin proteins are essential for caveola biogenesis and function. Here, we identify a role for the muscle-specific component, Cavin4, in skeletal muscle T-tubule development by analyzing two vertebrate systems: mouse and zebrafish. In both models Cavin4 localized to T-tubules and loss of Cavin4 resulted in aberrant T-tubule maturation. In zebrafish, which possess duplicatedcavin4paralogs, Cavin4b was shown to directly interact with the T-tubule-associated BAR domain protein, Bin1. Loss of both Cavin4a and Cavin4b caused aberrant accumulation of interconnected caveolae within the T-tubules, a fragmented T-tubule network enriched in Caveolin-3, and an impaired Ca2+response upon mechanical stimulation. We propose a role for Cavin4 in remodeling the T-tubule membrane early in development by recycling caveolar components from the T-tubule to the sarcolemma. This generates a stable T-tubule domain lacking caveolae that is essential for T-tubule function.